mediator of BCR-ABL–driven leukemogenesis

Genetic loss-of-function studies in murine tumor models have been essential in the analysis of downstream mediators of onco-genic transformation. Unfortunately, these studies are frequently limited by the availability of genetically modified mouse strains. Here we describe a versatile method allowing the efficient expression of an oncogene and simultaneous knockdown of targets of interest (TOI) from a single retroviral vector. Both oncogene and TOI-specific miR30-based shRNA are under the control of the strong viral long terminal repeat promoter, resulting in a single shared RNA transcript. Using this vector in a mu-rine syngeneic BM transplantation model for BCR-ABL–induced chronic myeloid leu-kemia, we find that oncogene expression andtargetknockdowninprimaryhematopoi-etic cells with this vector is efficient both in vitro and in vivo, and demonstrate that Raf1, but not BRAF, modulates BCR-ABL–depen-dent ERK activation and transformation of hematopoietic cells. This expression sys-tem could facilitate genetic loss-of-function studies and allow the rapid validation of potential drug targets in a broad range of oncogene-driven murine tumor models. (Blood. 2011;118(8):2200-2210